Cancer cells from a primary focus metastasize all over the body through blood vessels and lymphatic vessels. Because the primary focus is removed as much as possible in cancer surgery, the accurate detection of metastasis and the appropriate treatment depending on the degree of metastasis are necessary. Therefore, diagnosing lymph node metastasis by cancer cells is extremely important for selecting the appropriate treatment for cancer.
The diagnosis of lymph node metastasis by cancer cells is broadly divided into pretreatment diagnostic imaging and posttreatment (postoperative) pathological diagnosis. The diagnostic imaging methods that are used for detecting lymph node metastasis in cancer (inspection for the presence of lymph node metastasis) include computed tomography (CT); positron emission tomography (PET); PET-CT, which uses an apparatus integrating PET and CT; and endoscopic ultrasoundscopy (EUS); however, the diagnostic imaging methods have difficulty or only limited availability in detecting microscopic lymph node metastasis. On the other hand, pathological diagnosis method utilizes specimens prepared from a number of excised lymph node tissues under a microscope and is a highly accurate and reliable diagnostic method; however, the diagnosis can only be made using excised lymph nodes as posttreatment (postoperative) diagnosis and therefore cannot be used for selecting the optimum treatment in advance. The diagnosis of lymph node metastasis by cancer cells is problematic in that the pretreatment diagnosis depends on the diagnostic imaging, which is currently less accurate, whereas reliable diagnosis is made in the posttreatment (postoperative) pathological diagnosis.
Therefore, molecular diagnostic techniques using molecular markers are important in the diagnosis of lymph node metastasis by cancer cells, and several techniques have been developed. Many conventionally known molecular diagnostic techniques utilize a protein (target protein) that is not expressed or expressed at a lower level in normal cells and is highly expressed in cancer cells or a nucleic acid (target nucleic acid, as a general term for DNA, mRNA, cDNA, etc.) included in a gene encoding the target protein. Specifically, a target protein included in lymph node tissues resected/excised from a living body is detected using an immunoassay, or conversely, a target nucleic acid is amplified using loop-mediated isothermal amplification (LAMP) or polymerase chain reaction (PCR) to detect the amplification product using a known method to determine the presence of metastatic cancer cells.
Regarding molecular diagnostic techniques, for example, Patent Literature 1 (Japanese Laid-Open Patent Publication No. 2007-175021) proposed a method for determining the presence of lymph node metastasis by colon cancer cells using the mRNA or a fragment of a gene encoding at least one protein selected from the group consisting of PIGR, CLDN3, LGALS4, AGR2, TACSTD1, GPX2, RAI3, TSPAN1, CKB, ELF3, FXYD3, CDH1, REG4, GDF 15, CLDN4, OLFM 4, CD9, CDH17, SELENBP, LCN2, TMPRSS4, CFTR, TM4SF3, ID1, CYP2S1, TFF3, EHF, FAT, KLF5, SLC9A3R2, HOXB9, ATP1B1, PCK1, and FCGBP. Patent Literature 2 (Japanese Laid-Open Patent Publication No. 2007-037421) described the determination of lymph node metastasis in colon cancer by entering the value of expression of a gene set represented by the database access numbers (serial numbers) NM_003404 (G1592), NM_002128 (G2645), NM_052868 (G3031), NM_005034 (03177), NM_001540 (G3753), NM_005722 (G3826), and NM_015315(G4370) into a mathematical function. Patent Literature 3 (Japanese Laid-Open Patent Publication No. 2008-020438) describes that lymph node metastasis, e.g., from breast cancer, can be determined with higher reliability by determining the expression of a polypeptide related to cytokeratin in a sample prepared from lymph node tissue.
On the other hand, it was recently determined that inflammatory responses promote carcinogenesis by damaging DNA, stimulating angiogenesis and cell proliferation, and inhibiting apoptosis. In this regard, serum C-reactive protein (CRP) has been investigated as a risk factor and a prognostic factor in colon (Non Patent Literature 1: Erlinger T. P. et al., JAMA 2004; 291; 585-590), esophageal (Non Patent Literature 2: Shimada H. et al., J. Surg. Oncol. 2003; 83; 248-252), hepatocellular (Non Patent Literature 3: Hashimoto K. et al., Cancer 2005; 103; 1856-1864), renal (Non Patent Literature 4: Miyata Y. et al, Urology 2001; 58; 161-164), and ovarian (Non Patent Literature 5: Hefler L. A. et al., Clin. Cancer Res. 2008; 14; 710-714) cancers.
A higher serum CRP level is considered to be associated with a higher risk of developing cancer. For example, Non Patent Literature 6 (Nozoe T. et al., Am. J. Surg. 1998; 176(4):335-8) describes that liver metastasis and lymph node metastasis in colon cancer patients are associated with preoperative increases in serum CRP levels, Non Patent Literature 7 (Nozoe T. et al., Am. J. Surg. 2001; 182(2), 197-201) describes that lymph node metastasis in esophageal cancer patients is associated with preoperative increases in serum CRP levels, and Non Patent Literature 8 (Ines G. et al., World J. Gastroenterol. 2006; 12(23), 3746-3750) describes that a higher serum CRP level in esophageal cancer patients is associated with lymph node metastasis.
It has been reported that genetic polymorphisms are strongly related to serum CRP levels (Non Patent Literature 9: Carlson C. S. et al., Am. J. Hum. Gen. 2005; 77; 64-77 and Non Patent Literature 10: Szalai A. J. et al., J. Mol. Med. 2005; 83; 440-447).
Therefore, the present inventors examined whether CRP genetic polymorphisms act as cancer progression factors in esophageal cancer patients. Although we consequently revealed the potential association of CRP-717T>C genetic polymorphisms with lymph node metastasis (Non Patent Literature 11: Motoyama et al., The Japanese Journal of Gastroenterological Surgery, vol. 41, No. 7, pp. 1169, July 2008), a technique for detecting lymph node metastasis by cancer cells using the CRP-717T>C genetic polymorphism suffered from lower determination accuracy and was not put into practical use because metastasis was not statistically significant.